The Putative Cellodextrin Transporter-like Protein CLP1 Is Involved in Cellulase Induction in Neurospora crassa

Cai, Pengli; Wang, Bang; Ji, Jingxiao; Jiang, Yongsheng; Wan, Li; Tian, Chaoguang; Ma, Yanhe

doi:10.1074/jbc.m114.609875

Cited by 36 publications

(31 citation statements)

References 30 publications

(41 reference statements)

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“…CDT-1 and CDT-2 are two important cellodextrin transporters used by N. crassa in cellulose degradation [25], of which only CDT-2 is also involved in hemicellulose utilization [7,26]. In previous results, the discovery of the xylodextrin consumption pathway consisting of transporter CDT-2 along with two β-xylosidases GH43-2 and GH43-7 provided new modes for utilization of xylose derived from hemicellulose [7].…”

Section: Discussionmentioning

confidence: 99%

Screening and directed evolution of transporters to improve xylodextrin utilization in the yeast Saccharomyces cerevisiae

Zhang

Acosta-Sampson

et al. 2017

Preprint

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The economic production of cellulosic biofuel requires efficient and full utilization of all abundant carbohydrates naturally released from plant biomass by enzyme cocktails.Recently, we reconstituted the Neurospora crassa xylodextrin transport and consumption system in Saccharomyces cerevisiae, enabling growth of yeast on xylodextrins aerobically. However, the consumption rate of xylodextrin requires improvement for industrial applications, including consumption in anaerobic conditions. As a first step in this improvement, we report analysis of orthologues of the N. crassa transporters CDT-1 and CDT-2. Transporter ST16 from Trichoderma virens enables faster aerobic growth of S. cerevisiae on xylodextrins compared to CDT-2. ST16 is a xylodextrin-specific transporter, and the xylobiose transport activity of ST16 is not inhibited by cellobiose. Other transporters identified in the screen also enable growth on xylodextrins including xylotriose. Taken together, these results indicate that multiple transporters might prove useful to improve xylodextrin utilization in S. cerevisiae. Efforts to use directed evolution to improve ST16 from a chromosomally-integrated copy were not successful, due to background growth of yeast on other carbon sources present in the selection medium. Future experiments will require increasing the baseline growth rate of the yeast population on xylodextrins, to ensure that the selective pressure exerted on xylodextrin transport can lead to isolation of improved xylodextrin transporters.

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Section: Discussionmentioning

confidence: 99%

Screening and directed evolution of transporters to improve xylodextrin utilization in the yeast Saccharomyces cerevisiae

Zhang

Acosta-Sampson

et al. 2017

Preprint

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“…Though a number of papers on the fundamental research of the cellulase derived from N. crassa have been published, this is the first report on the heterologous expression and production of the cellulase component CBHI (NCU07340) in P. pastoris . There were extensive and intensive researches on the heterologous expression and production of T. reesei cellulase components in P. pastoris because T. reesei cellulase had wide applications in industry .…”

Section: Discussionmentioning

confidence: 99%

Heterologous expression of Neurospora crassa cbh1 gene in Pichia pastoris resulted in production of a neutral cellobiohydrolase I

Yang

Deng

Zhao

et al. 2019

Biotechnology Progress

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The high production cost of cellulase is one of the limitations that hinder the commercialization of lignocellulose-based biorefineries. As one of the important cellulases, Neurospora crassa cellulase is not so intensively investigated as T. reesei cellulase.In this study, the cbh1gene (NCU07340) cloned from N. crassa was expressed in Pichia pastoris under the control of alcohol oxidase 1 (AOX1) promotor. Six transformants with the highest resistance to G418 were selected by two rounds of transformant screening, among which the most robust producer of the recombinant cellobiohydrolase I (CBHI) has an Avicelase activity of 0.22 U/mL. After fermentation optimization, it was improved to 0.30 U/mL. Interestingly, the optimal temperature and pH of the recombinant CBHI were 60 C and 7.2, respectively, and it was quite stable within the wide ranges of temperature and pH. This work is a good example for the future improvement and optimization of N. crassa cellulase.

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“…Although CLP1 protein cannot transport cellodextrin, this signaling sensor may suppress cellulase induction. The co-disruption of cdt2 and clp1 enhanced 6.9-fold the cellulase production with cellobiose induction in the strain ∆3βG [20]. In P. echinulatum 2HH, clp1 was identified as orthologous to PECH 007291, while cdt1 is orthologous to PECH 005610 and cdt2 is orthologous to PECH 004467.…”

Section: Sugar Transporters (Sts)mentioning

confidence: 99%

“…This pathway acts when cellobiose or other cellodextrins accumulates into the cell, raising the secretion of cellulases. In this context, intracellular accumulation of cellodextrins can occur in two ways: i) by low activity of iBGLs, which reduces the hydrolysis of cellodextrins to D-glucose [19,34]; and ii) by the high expression of STs, which are able to transport cellodextrins from the medium into the cell [20,35]. In order to discover the features behind this cellodextrin induction system in P. echinulatum 2HH, we performed several genomic analyses including iBGLs and STs, as presented below.…”

Section: Cellulolytic System Expression Induced By Cellodextrinsmentioning

confidence: 99%

“…Along these lines, it has been shown that cellulolytic enzyme expression is induced by cellobiose in many species of fungi, regarding cellobiose is the primary end product generated from cellulose degradation by cellulases [18]. Recent research results support that accumulation of intracellular cellodextrins (mainly cellobiose) may raise cellulases secretion by a cascade signaling pathway in P. oxalicum [19] and Neurospora crassa [20].…”

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confidence: 99%

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CAZyome and Sugar Transportome Unveil Singular Aspects of the Lignocellulolytic Enzyme System of Penicillium echinulatum 2HH

Lenz

Balbinot

Oliveira

et al. 2020

Preprint

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Background: The production of bioethanol using lignocellulosic feedstocks has proved to be a cutting-edge technology. However, this technology faces limitations such as cost and yield of enzymatic systems, required to degrade efficiently the polysaccharides of plant cell walls. Penicillium echinulatum 2HH is a widely studied ascomycete, known by its efficient cellulolytic cocktails. One strategy to improve the saccharification yields for commercial exploitation is the design of hypersecreting strains. However, molecular knowledge about the lignocellulolytic system of this specific fungus is scarce. Understanding both lignocellulolytic and sugar uptake systems is essential to obtain industrial strains with adequate efficiency for bioethanol production. Results: We report a comprehensive in silico characterization of CAZymes and sugar transporters of P. echinulatum 2HH. The CAZyome reveals an outstanding repertoire of enzymes involved in plant biomass degradation. Among them, we highlight the cellulolytic enzyme system whose genes are predominantly orthologous to P. oxalicum 114-2, demonstrating the high similarity of these phylogenetically related enzyme producers. We also report a LPMO-type enzyme of the AA16 family described for the first time in these fungi. In addition to the well-known high activity of ß-glucosidases, we found that coding genes for the AA16, GH5-4 and GH45 families comprehend the main differences in the cellulolytic complexes of P. echinulatum 2HH and P. oxalicum 114-2, when both are compared to commercial producers. Our phylogenetic analysis of the sugar transportome suggests that P. echinulatum 2HH diversity and specificity of STs include eight major families with specificity to different groups of sugars. Finally, our phylogenetic analyses enabled the identification of several iBGLs and STs potentially involved in the accumulation of intracellular cellodextrins.Conclusions: Overall, both CAZyome and sugar transportome of P. echinulatum revealed new insights into the mechanisms underlying a flexible and highly functional metabolism to degrade plant biomass. Peculiarities found in our study help to highlight the cellulolytic complex of P. echinulatum 2HH, contributing to the commercial ascendance of Penicillium spp. as cellulolytic enzyme producers. Furthermore, the first phylogenetic classification of STs and iBGLs shed new light into the role of these genes regarding the preferred carbon source during fungal growth. Along these lines, these iBGLs and STs comprise valuable gene targets to understand the regulatory mechanisms underlying cellulolytic enzymes and to design hypersecreting strains with adequate efficiency for bioethanol production in large-scale.

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The Putative Cellodextrin Transporter-like Protein CLP1 Is Involved in Cellulase Induction in Neurospora crassa

Cited by 36 publications

References 30 publications

Screening and directed evolution of transporters to improve xylodextrin utilization in the yeast Saccharomyces cerevisiae

Screening and directed evolution of transporters to improve xylodextrin utilization in the yeast Saccharomyces cerevisiae

Heterologous expression of Neurospora crassa cbh1 gene in Pichia pastoris resulted in production of a neutral cellobiohydrolase I

CAZyome and Sugar Transportome Unveil Singular Aspects of the Lignocellulolytic Enzyme System of Penicillium echinulatum 2HH

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